LOCUS       sequence-205290-        5858 bp    DNA     linear   UNA 15-APR-2025
DEFINITION  Expresses PseC from C. jejuni with an N-terminal octaHis tag and TEV
            cleavage site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5858)
  AUTHORS   Zamora CY, Schocker NS, Chang MM, Imperiali B
  TITLE     Chemoenzymatic Synthesis and Applications of Prokaryote-Specific 
            UDP-Sugars.
  JOURNAL   Methods Enzymol. 2017;597:145-186. doi: 10.1016/bs.mie.2017.06.003. 
            Epub 2017 Jul 5.
  PUBMED    28935101
REFERENCE   2  (bases 1 to 5858)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Methods 
            Enzymol."; date: "2017"; volume: "597:145-186. doi"; pages: " 
            10.1016/bs.mie.2017.06.003. Epub 2017 Jul 5"
COMMENT     Sequence Label: His8-TEV-PseC
FEATURES             Location/Qualifiers
     source          1..5858
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             116..136
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1280..1297
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1350..1368)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1364..1411
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1448..1903
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1535..1554)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1745..1766
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1995..2810)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1995..2810)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2718..2737
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2802..2821
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2932..3520
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3421..3440
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3674..3691
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3706..3845
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3771..3790)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3931..3953
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4386..5468)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5430..5449
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5469..5546)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5752..5771
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gcaccatcag catcatcatc atcacgaaaa
      121 tctgtatttt cagagcggat ccatgattac ttattctcat caaaatattg atcaaagcga
      181 tatagatgcg cttacaaaag ctttaaaaga tgaaatttta acagggggta aaaaggtaga
      241 tgaatttgaa gaagcacttt gcgaatacat aggtgttaag cacgcttgtg tgttaaattc
      301 ggctacttct gcccttcatc ttgcctatac agcactaggc atcaaagaaa aaatcgtttt
      361 aaccactcct ttaaccttcg ctgctacggc taatgcggcc ttaatagctg gagctaaggt
      421 agagtttatc gacattaaaa atgatggcaa tatagatgag aaaaagctcg aagcaaggct
      481 tgtaaaaaat agcaccgata taggagctat tagcgtggtg gattttggag gaaatagtgt
      541 tgaaatggat gagatttcaa atcttgctaa aaaatacaat atccctttaa tcgatgatgc
      601 aagccatgct ttaggtagcg agtataaggg taaaaaagtg ggttctatgg cggatttaag
      661 tattttttct tttcatcctg ttaagcctat taccactttt gaggggggtg ctgtagtaag
      721 caataataaa gaattaatct ccaaaataaa acttttaaga agccatggta tcgttaaaaa
      781 aaggctttgg gatagcgata tgatagaact tggctataat taccgcttaa gtgatgttgc
      841 ttgtgcttta gggataaacc agcttaaaaa actcgatcac aatcttgaaa aacgagaaga
      901 aatcacaagt ttttacgata aagaatttga aaaaaatcat tatttttcca cgataaaaat
      961 caaagactat aaaaaaagct caaggcattt ataccctatc ttgctttttc ctgaatttta
     1021 ttgtcaaaaa gaagaacttt ttgaaagctt gcttcatgca ggaattggcg tacaggtaca
     1081 ttataaacca acttatgagt ttagttttta taaaaaactc ttaggagaaa tcagactcca
     1141 aaatgcagat aatttttaca aagccgagct tagcatacct tgtcatcaag aaatgaatct
     1201 aaaagatgca aaatttgtaa aagatacttt atttagcatt cttgaaaaag ttaaaaaggg
     1261 atattgtgga taactcgagc accaccacca ccaccactga gatccggctg ctaacaaagc
     1321 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
     1381 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc
     1441 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     1501 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     1561 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     1621 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     1681 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     1741 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     1801 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     1861 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
     1921 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     1981 tcatgaatta attcttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata
     2041 tcaggattat caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca
     2101 ccgaggcagt tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca
     2161 acatcaatac aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca
     2221 ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact
     2281 tgttcaacag gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta
     2341 ttcattcgtg attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta
     2401 caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca
     2461 cctgaatcag gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg
     2521 agtaaccatg catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat
     2581 tccgtcagcc agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg
     2641 ccatgtttca gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca
     2701 cctgattgcc cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg
     2761 gaatttaatc gcggcctaga gcaagacgtt tcccgttgaa tatggctcat aacacccctt
     2821 gtattactgt ttatgtaagc agacagtttt attgttcatg accaaaatcc cttaacgtga
     2881 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     2941 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     3001 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     3061 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     3121 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     3181 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     3241 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     3301 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     3361 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     3421 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     3481 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     3541 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     3601 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     3661 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt
     3721 tctccttacg catctgtgcg gtatttcaca ccgcaatggt gcactctcag tacaatctgc
     3781 tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac tgggtcatgg
     3841 ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg
     3901 catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac
     3961 cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg
     4021 attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta
     4081 atgtctggct tctgataaag cgggccatgt taaggggagc gagggcgtgc aagattccga
     4141 ataccgcaag cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa
     4201 tgacccagag cgctgccggc acctgtccta cgagttgcat gataaagaag acagtcataa
     4261 gtgcggcgac gatagtcatg ccccgcgccc accggaagga gctgactggg ttgaaggctc
     4321 tcaagggcat cggtcgagat cccggtgcct aatgagtgag ctaacttaca ttaattgcgt
     4381 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     4441 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc
     4501 agtgagacgg gcaacagctg attgcccttc accgcctggc cctgagagag ttgcagcaag
     4561 cggtccacgc tggtttgccc cagcaggcga aaatcctgtt tgatggtggt taacggcggg
     4621 atataacatg agctgtcttc ggtatcgtcg tatcccacta ccgagatgtc cgcaccaacg
     4681 cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc
     4741 agcatcgcag tgggaacgat gccctcattc agcatttgca tggtttgttg aaaaccggac
     4801 atggcactcc agtcgccttc ccgttccgct atcggctgaa tttgattgcg agtgagatat
     4861 ttatgccagc cagccagacg cagacgcgcc gagacagaac ttaatgggcc cgctaacagc
     4921 gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca
     4981 tgggagaaaa taatactgtt gatgggtgtc tggtcagaga catcaagaaa taacgccgga
     5041 acattagtgc aggcagcttc cacagcaatg gcatcctggt catccagcgg atagttaatg
     5101 atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg
     5161 ccgcttcgtt ctaccatcga caccaccacg ctggcaccca gttgatcggc gcgagattta
     5221 atcgccgcga caatttgcga cggcgcgtgc agggccagac tggaggtggc aacgccaatc
     5281 agcaacgact gtttgcccgc cagttgttgt gccacgcggt tgggaatgta attcagctcc
     5341 gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc
     5401 acgcgggaaa cggtctgata agagacaccg gcatactctg cgacatcgta taacgttact
     5461 ggtttcacat tcaccaccct gaattgactc tcttccgggc gctatcatgc cataccgcga
     5521 aaggttttgc gccattcgat ggtgtccggg atctcgacgc tctcccttat gcgactcctg
     5581 cattaggaag cagcccagta gtaggttgag gccgttgagc accgccgccg caaggaatgg
     5641 tgcatgcaag gagatggcgc ccaacagtcc cccggccacg gggcctgcca ccatacccac
     5701 gccgaaacaa gcgctcatga gcccgaagtg gcgagcccga tcttccccat cggtgatgtc
     5761 ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc
     5821 ggcgtagagg atcgagatcg atctcgatcc cgcgaaat
//
